Poly(lactate-co-glycolate) (PLGA), a random copolymer of lactate and glycolate, is a representative biodegradable polymer which is highly applicable as a general-purpose polymer or a medical polymer. Currently, PLGA can be produced by the direct polymerization reaction of lactate with glycolate, but this reaction mainly produces a PLGA having only a low molecular weight (1,000-5,000 Daltons). PLGA having a high molecular weight of 100,000 Daltons or more can be synthesized by the ring-opening polymerization of lactide with glycolide. Lactide and glycolide are cyclic diesters of lactate and glycolate, respectively, and are produced by thermal decomposition of a lactate oligomer and a glycolate oligomer, respectively. The ring-opening polymerization requires the use of a catalyst such as tin(II) 2-ethylhexanoate, tin(II) alkoxide, aluminum isopropoxide or the like. There is a method of producing a higher-molecular-weight polymer from a low-molecular-weight polymer, obtained by direct polymerization, by use of a chain-coupling agent. However, since the chain-coupling agent is used, the method of producing high-molecular weight PLGA has disadvantages in that the addition of an organic solvent or a chain coupling agent makes the process complex and in that this organic solvent or chain-coupling agent is not easily removed. Current commercial processes for producing high-molecular-weight PLGA use a method that comprises converting lactate and glycolate into lactide and glycolide, respectively, and then synthesizing PLGA by the ring-opening polymerization of lactide with glycolide.
Meanwhile, polyhydroxyalkanoate (PHA) is a polyester which is produced when microorganisms accumulate excess carbon sources as energy or carbon source-storing substances in the cells when subjected to an environment deficient in nutrients such as phosphorus, nitrogen, magnesium, oxygen and the like. Since PHA is completely biodegradable and may have physical properties similar to those of conventional synthetic polymers produced from petroleum, it is attracting attention as an environmentally friendly substitute for petroleum-based synthetic plastics.
It is known that PHA can be produced by a variety of organisms, including Ralstonia eutropha, Pseudomonas, Bacillus, recombinant E. coli, and the like, and may contain about 150 or more different monomers. Such PHA is roughly divided into SCL-PHA (short-chain-length PHA) having a short chain length monomer (3-5 carbon atoms) and MCL-PHA (medium-chain-length PHA) having a longer chain length monomer. PHA synthases, which are key enzymes that synthesize PHA, are roughly divided into four classes by the kind of monomer, which is used as a substrate, and the subunits of the enzyme (Qi et al., FEMS Microbiol. Lett., 157:155, 1997; Qi et al., FEMS Microbiol. Lett., 167:89, 1998; Langenbach et al., FEMS Microbiol. Lett., 150:303, 1997; WO 01/55436; U.S. Pat. No. 6,143,952; WO 98/54329; WO 99/61624).
Glycolic acid is the simplest hydroxycarboxylic acid having two carbon atoms, and PHA that naturally contains glycolic acid has not been reported yet. However, glycolic acid together with polylactate is highly useful as a representative synthetic biopolymer, and thus various attempts have been made to insert it into a PHA monomer.
In a previous patent (U.S. Pat. No. 8,883,463 B2), the present inventors constructed an E. coli strain, which produces glycolate by using glucose as a carbon source without having to add an external precursor, by engineering the glyoxylate shunt pathway, and found that PLGA was produced from glucose by culturing the E. coli strain transformed with a gene encoding Clostridium propionicum-derived propionyl-CoA transferase (Pct), which is an enzyme that converts lactate and glycolate into lactyl-CoA and glycolyl-CoA, respectively, and a gene encoding polyhydroxyalkanoate (PHA) synthase (PhaC1) which can use lactyl-CoA and glycolyl-CoA as substrates.
Accordingly, the present inventors have made extensive efforts to develop a recombinant E. coli strain capable of producing a PLGA having a high content of a glycolate fraction with higher efficiency by using xylose as a main carbon source without having to add glycolate from an external source. As a result, the present inventors have constructed a recombinant microorganism expressing Clostridium propionicum-derived propionyl-CoA transferase, Pseudomonas sp. 6-19-derived PHA synthase, xylose dehydrogenase and xylonolactonase, and have found that the recombinant microorganism produces poly(lactate-co-glycolate) and its copolymers by using xylose as a single carbon source or using xylose and glucose simultaneously as carbon sources, thereby completing the present invention.